Jiongjiong Chen

The draft genome of sweet orange (Citrus sinensis)

Oranges are an important nutritional source for human health and have immense economic value. Here we present a comprehensive analysis of the draft genome of sweet orange (Citrus sinensis). The assembled sequence covers 87.3% of the estimated orange genome, which is relatively compact, as 20% is composed of repetitive elements. We predicted 29,445 protein-coding genes, half of which are in the heterozygous state. With additional sequencing of two more citrus species and comparative analyses of seven citrus genomes, we present evidence to suggest that sweet orange originated from a backcross hybrid between pummelo and mandarin. Focused analysis on genes involved in vitamin C metabolism showed that GalUR, encoding the rate-limiting enzyme of the galacturonate pathway, is significantly upregulated in orange fruit, and the recent expansion of this gene family may provide a genomic basis. This draft genome represents a valuable resource for understanding and improving many important citrus traits in the future.

Miniature Inverted–Repeat Transposable Elements (MITEs) Have Been Accumulated through Amplification Bursts and Play Important Roles in Gene Expression and Species Diversity in Oryza sativa

Miniature inverted–repeat transposable elements (MITEs) are predicted to play important roles on genome evolution. We developed a BLASTN-based approach for de novo identification of MITEs and systematically analyzed MITEs in rice genome. The genome of rice cultivar Nipponbare (Oryza sativa ssp. japonica) harbors 178,533 MITE-related sequences classified into 338 families. Pairwise nucleotide diversity and phylogenetic tree analysis indicated that individual MITE families were resulted from one or multiple rounds of amplification bursts. The timing of amplification burst varied considerably between different MITE families or subfamilies. MITEs are associated with 23,623 (58.2%) genes in rice genome. At least 7,887 MITEs are transcribed and more than 3,463 were transcribed with rice genes. The MITE sequences transcribed with rice coding genes form 1,130 pairs of potential natural sense/antisense transcripts. MITEs generate 23.5% (183,837 of 781,885) of all small RNAs identified from rice. Some MITE families generated small RNAs mainly from the terminals, while other families generated small RNAs predominantly from the central region. More than half (51.8%) of the MITE-derived small RNAs were generated exclusively by MITEs located away from genes. Genome-wide analysis showed that genes associated with MITEs have significantly lower expression than genes away from MITEs. Approximately 14.8% of loci with full-length MITEs have presence/absence polymorphism between rice cultivars 93-11 (O. sativa ssp. indica) and Nipponbare. Considering that different sets of genes may be regulated by MITE-derived small RNAs in different genotypes, MITEs provide considerable diversity for O. sativa.

Dynamic Nucleotide-Binding-Site and Leucine-Rich-Repeat- encoding Genes in the Grass Family

The proper use of resistance genes (R genes) requires a comprehensive understanding of their genomics and evolution. We analyzed genes encoding nucleotide-binding sites and leucine-rich repeats in the genomes of rice (Oryza sativa), maize (Zea mays), sorghum (Sorghum bicolor), and Brachypodium distachyon. Frequent deletions and translocations of R genes generated prevalent presence/absence polymorphism between different accessions/species. The deletions were caused by unequal crossover, homologous repair, nonhomologous repair, or other unknown mechanisms. R gene loci identified from different genomes were mapped onto the chromosomes of rice cv Nipponbare using comparative genomics, resulting in an integrated map of 495 R loci. Sequence analysis of R genes from the partially sequenced genomes of an African rice cultivar and 10 wild accessions suggested that there are many additional R gene lineages in the AA genome of Oryza. The R genes with chimeric structures (termed type I R genes) are diverse in different rice accessions but only account for 5.8% of all R genes in the Nipponbare genome. In contrast, the vast majority of R genes in the rice genome are type II R genes, which are highly conserved in different accessions. Surprisingly, pseudogene-causing mutations in some type II lineages are often conserved, indicating that their conservations were not due to their functions. Functional R genes cloned from rice so far have more type II R genes than type I R genes, but type I R genes are predicted to contribute considerable diversity in wild species. Type I R genes tend to reduce the microsynteny of their flanking regions significantly more than type II R genes, and their flanking regions have slightly but significantly lower G/C content than those of type II R genes.

P-MITE: a database for plant miniature inverted-repeat transposable elements

Miniature inverted-repeat transposable elements (MITEs) are prevalent in eukaryotic species including plants. MITE families vary dramatically and usually cannot be identified based on homology. In this study, we de novo identified MITEs from 41 plant species, using computer programs MITE Digger, MITE-Hunter and/or Repetitive Sequence with Precise Boundaries (RSPB). MITEs were found in all, but one (Cyanidioschyzon merolae), species. Combined with the MITEs identified previously from the rice genome, >2.3 million sequences from 3527 MITE families were obtained from 41 plant species. In general, higher plants contain more MITEs than lower plants, with a few exceptions such as papaya, with only 538 elements. The largest number of MITEs is found in apple, with 237 302 MITE sequences. The number of MITE sequences in a genome is significantly correlated with genome size. A series of databases (plant MITE databases, P-MITE), available online at http://pmite.hzau.edu.cn/django/mite/, was constructed to host all MITE sequences from the 41 plant genomes. The databases are available for sequence similarity searches (BLASTN), and MITE sequences can be downloaded by family or by genome. The databases can be used to study the origin and amplification of MITEs, MITE-derived small RNAs and roles of MITEs on gene and genome evolution.

Frequent loss of lineages and deficient duplications accounted for low copy number of disease resistance genes in Cucurbitaceae

BackgroundThe sequenced genomes of cucumber, melon and watermelon have relatively few R-genes, with 70, 75 and 55 copies only, respectively. The mechanism for low copy number of R-genes in Cucurbitaceae genomes remains unknown.ResultsManual annotation of R-genes in the sequenced genomes of Cucurbitaceae species showed that approximately half of them are pseudogenes. Comparative analysis of R-genes showed frequent loss of R-gene loci in different Cucurbitaceae species. Phylogenetic analysis, data mining and PCR cloning using degenerate primers indicated that Cucurbitaceae has limited number of R-gene lineages (subfamilies). Comparison between R-genes from Cucurbitaceae and those from poplar and soybean suggested frequent loss of R-gene lineages in Cucurbitaceae. Furthermore, the average number of R-genes per lineage in Cucurbitaceae species is approximately 1/3 that in soybean or poplar. Therefore, both loss of lineages and deficient duplications in extant lineages accounted for the low copy number of R-genes in Cucurbitaceae. No extensive chimeras of R-genes were found in any of the sequenced Cucurbitaceae genomes. Nevertheless, one lineage of R-genes from Trichosanthes kirilowii, a wild Cucurbitaceae species, exhibits chimeric structures caused by gene conversions, and may contain a large number of distinct R-genes in natural populations.ConclusionsCucurbitaceae species have limited number of R-gene lineages and each genome harbors relatively few R-genes. The scarcity of R-genes in Cucurbitaceae species was due to frequent loss of R-gene lineages and infrequent duplications in extant lineages. The evolutionary mechanisms for large variation of copy number of R-genes in different plant species were discussed.

Dramatic Number Variation of R Genes in Solanaceae Species Accounted for by a Few R Gene Subfamilies.

Most disease resistance genes encode nucleotide-binding-site (NBS) and leucine-rich-repeat (LRR) domains, and the NBS-LRR encoding genes are often referred to as R genes. Using newly developed approach, 478, 485, 1,194, 1,665, 2,042 and 374 R genes were identified from the genomes of tomato Heinz1706, wild tomato LA716, potato DM1-3, pepper Zunla-1 and wild pepper Chiltepin and tobacco TN90, respectively. The majority of R genes from Solanaceae were grouped into 87 subfamilies, including 16 TIR-NBS-LRR (TNL) and 71 non-TNL subfamilies. Each subfamily was annotated manually, including identification of intron/exon structure and intron phase. Interestingly, TNL subfamilies have similar intron phase patterns, while the non-TNL subfamilies have diverse intron phase due to frequent gain of introns. Prevalent presence/absence polymorphic R gene loci were found among Solanaceae species, and an integrated map with 427 R loci was constructed. The pepper genome (2,042 in Chiltepin) has at least four times of R genes as in tomato (478 in Heinz1706). The high number of R genes in pepper genome is due to the amplification of R genes in a few subfamilies, such as the Rpi-blb2 and BS2 subfamilies. The mechanism underlying the variation of R gene number among different plant genomes is discussed.

Miniature inverted-repeat transposable elements (MITEs) in rice were originated and amplified predominantly after the divergence of Oryza and Brachypodium and contributed considerable diversity to the species

Miniature inverted-repeat transposable elements (MITEs) are ubiquitous in high eukaryotic genomes. More than 178,000 MITE sequences of 338 families are present in the genome of rice (Oryza sativa) cultivar Nipponbare. Interestingly, only two of the 338 MITE families have homologous sequences in the genome of Brachypodium distachyon, a relative in the grass family. Therefore, the vast majority of MITEs in the rice genome were originated and amplified after the divergence of Oryza and Brachypodium. Comparison between rice cultivar Nipponbare and another rice cultivar 93–11 showed 14.8% of MITEs exhibit presence/absence (P/A) polymorphism. The P/A polymorphism was mainly attributed to recent MITE transpositions, while less than 10% of the P/A polymorphism was caused by MITE excisions. Therefore, the high P/A polymorphisms of MITEs may generate considerable gene expression and phenotypic diversity for O. sativa.

The I2 resistance gene homologues in Solanum have complex evolutionary patterns and are targeted by miRNAs

BackgroundSeveral resistance traits, including the I2 resistance against tomato fusarium wilt, were mapped to the long arm of chromosome 11 of Solanum. However, the structure and evolution of this locus remain poorly understood.ResultsComparative analysis showed that the structure and evolutionary patterns of the I2 locus vary considerably between potato and tomato. The I2 homologues from different Solanaceae species usually do not have orthologous relationship, due to duplication, deletion and frequent sequence exchanges. At least 154 sequence exchanges were detected among 76 tomato I2 homologues, but sequence exchanges between I2 homologues in potato is less frequent. Previous study showed that I2 homologues in potato were targeted by miR482. However, our data showed that I2 homologues in tomato were targeted by miR6024 rather than miR482. Furthermore, miR6024 triggers phasiRNAs from I2 homologues in tomato. Sequence analysis showed that miR6024 was originated after the divergence of Solanaceae. We hypothesized that miR6024 and miR482 might have facilitated the expansion of the I2 family in Solanaceae species, since they can minimize their potential toxic effects by down-regulating their expression.ConclusionsThe I2 locus represents a most divergent resistance gene cluster in Solanum. Its high divergence was partly due to frequent sequence exchanges between homologues. We propose that the successful expansion of I2 homologues in Solanum was at least partially attributed to miRNA mediated regulation.

Evolutionary Genomics of Miniature Inverted-Repeat Transposable Elements (MITEs) in Plants

More and more evidence has accumulated in the past 20 years suggesting that MITEs may have played important roles in plant gene and genome evolution. With a large number of plant genomes sequenced and the development of computational programs for de novo MITE identification, a massive number of MITEs have been identified from plant genomes. The number of MITEs in a genome varied dramatically among different plant species. There is significant correlation between the number of MITEs and genome size, though there are several prominent exceptions. Some MITE families have a high copy number in a genome, probably due to one or several rounds of amplification bursts. Different MITE families in the same genome may have experienced amplification burst at different times, suggesting that their amplifications were triggered by distinct environments (such as stress) or genetic events. However, very few MITEs in plant genomes are currently active. MITEs are often distributed in gene-rich regions, and may be inserted in genes’ promoter regions or transcribed regions. They may affect (either upregulate or downregulate) the expression of nearby genes. MITEs may downregulate genes through small RNAs, which may be produced via NAT or double-stranded RNAs formed by transcribed MITE sequences. The presence/absence of MITEs as well as their potential effects on expression of nearby genes suggests that MITE may provide considerable physiological and phenotypic variations for a species. Important future studies on MITEs include the mechanisms of MITE activation and the effects of MITEs on gene and genome evolution.